Adult neural stem cells: plasticity and developmental potential.

Stem cells play an essential role during the processes of embryonic tissue formation and development and in the maintenance of tissue integrity and renewal throughout adulthood. The differentiation potential of stem cells in adult tissues has been thought to be limited to cell lineages present in the organ from which they derive, but there is evidence that somatic stem cells may display a broader differentiation repertoire. This has been documented for bone marrow stem cells (which can give rise to muscle, hepatic and brain cells) and for muscle precursors, which can turn into blood cells. The adult central nervous system (CNS) has long been considered incapable of cell renewal and structural remodeling. Recent findings indicate that, even in postnatal and adult mammals, neurogenesis does occur in different brain regions and that these regions actually contain adult stem cells. These cells can be expanded both in vivo and ex vivo by exposure to different combinations of growth factors and subsequently give rise to a differentiated progeny comprising the major cell types of the CNS. Almost paradoxically, adult neural stem cells display a multipotency much broader than expected, since they can differentiate into non-CNS mesodermal-derivatives, such as blood cells and skeletal muscle cells. We review the recent findings documenting this unforeseen plasticity and unexpected developmental potential of somatic stem cells in general and of neural stem cells in particular. To better introduce these concepts, some basic notions on the functional properties of adult neural stem cells will also be discussed, particularly focusing on the emerging role of the microenvironment in determining and maintaining their peculiar characteristics.     

Arachidonate release and c-fos expression in various models of hypoxia and hypoxia–hypoglycemia in retinoic acid differentiated neuroblastoma cells

Hypoxia-hypoglycemia has played an important role in inducing both phospholipase A2 activation and the expression of the early gene c-fos, in the neuroblastoma cell line SK-N-BE, after it has been differentiated by retinoic acid. Under hypoxic-hypoglycemic conditions, arachidonic acid release has found to be significant after 30 min, whereas c-fos expression has required at least 4 h. This model has been obtained by adding glycolytic inhibitor 2-deoxyglucose to the culture and by placing cells in an atmosphere containing 100% N2 for different time periods. This condition has been compared with two different models: NaCN and nitrogen have been used as hypoxic stimuli, without inhibiting the glycolytic pathway, but the same cell cultures have been used. Cell viability and the fall of cellular ATP levels have been evaluated in all the models, in order to monitor and compare the hypoxic cellular damage. Phospholipase A2 activation has been found to be significant in all conditions, even if to a different extent; but only hypoxia combined with the inhibition of the glycolytic pathway, has induced a significant expression of c-fos. It is very difficult to study hypoxic stimuli in 'in vitro' systems. Our study has compared three different models and the one combining gaseous hypoxia and hypoglycemic conditions seems to be very effective in stimulating early events involved in hypoxic phenomena such as phospholipase activation and the expression of the early gene c-fos.     

Enzyme-assisted photosensitization activates different apoptotic pathways in Rose Bengal acetate treated HeLa cells

Photosensitization of tumor cells after incubation with Rose Bengal acetate (RB-Ac) induces multiple organelle photodamage followed by apoptotic cell death. We used immunocytochemical techniques in multicolor fluorescence microscopy to elucidate whether this occurs through the simultaneous activation of different apoptotic pathways, in HeLa cells. We detected in situ the activated forms of caspases 9 and 3, and the translocation from the mitochondria to the nucleus of the apoptosis inducing factor; DNA electrophoretic techniques were also used to assess the occurrence of nuclear DNA cleavage into either high- or low-molecular-weight fragments. Both the caspase-dependent and caspase-independent apoptotic pathways are activated. The genomic DNA is degraded into high molecular weight molecules only, without the formation of oligonucleosome-sized fragments. The ability of RB-Ac to induce the simultaneous release of apoptogenic signals from different photodamaged organelles makes it an especially powerful cytotoxic agent.     

The Plasmodium falciparum protein Pfg27 is dispensable for gametocyte and gamete production, but contributes to cell integrity during gametocytogenesis

In the human malaria parasite Plasmodium falciparum , gametocyte maturation is a process remarkably longer than in other malaria species, accompanied by expression of 2–300 sexual stage-specific proteins. Disruption of several of their encoding genes so far showed that only the abundant protein Pfg27, produced at the onset of sexual differentiation, is essential for gametocyte production. In contrast with what has been previously described, here we show that P. falciparum pfg27 disruptant lines are able to undergo all stages of gametocyte maturation, and are able to mature into gametes. A fraction of Pfg27-defective gametocytes show, however, distinct abnormalities in intra- and extra-cellular membranous compartments, such as accumulation of parasitophorous vacuole-derived vesicles in the erythrocyte cytoplasm, large intracellular vacuoles and discontinuities in their trilaminar cell membrane. This work revises current knowledge on the role of Pfg27, indicating that the protein is not required for parasite entry into sexual differentiation, and suggesting that it is instead involved in maintaining cell integrity in the uniquely long gametocytogenesis of P. falciparum .     

Current status in production and utilization of dihaploids from somatic hybrids between eggplant (Solanum melongena L.) and its wild relatives

The major constrains for practical exploitation of the somatic hybrids between eggplant and its wild relatives have been their sterility and tetraploidy which prevented their incorporation into breeding programs. Here we demonstrate that anther culture was successfully utilized to bring back the ploidy level to the diploid status in tetraploid interspecific hybrids between eggplant and the allied species S. integrifolium and S. aethiopicum gr. gilo. Both the relative species are resistant to Fusarium oxysporum f. sp. melongenae and to some strains of bacterial wilt (Ralstonia solanacearum) which are very destructive diseases of eggplant. Dihaploid androgenetic plants were obtained from the somatic hybrids, from the “double somatic hybrid” obtained by sexual cross of the two somatic hybrids [(eggplant + S. aethiopicum) × (eggplant + S. integrifolium)], and from tetraploid backcrossed plants between the somatic hybrid with S. aethiopicum and eggplant. Phenotypical, molecular, biological and biochemical characterization, and also artificial inoculation with Fusarium oxysporum are consistent with a recombination between the genomes of the species involved in the hybridizations. Dihaploids resistant to Fusarium were successfully backcrossed with eggplant. Besides their utility as potential valuable breeding materials, the introgressed lines obtained may be utilized in genetic and molecular studies about the resistance to Fusarium from S. integrifolium and S. aethiopicum gr. gilo.     

Description of Turritopsoides marhei sp. nov. (Hydrozoa,Anthoathecata) from the Maldives and its phylogenetic position

Turritopsoides marhei, a new species of the hydrozoan family Oceaniidae, is described from the Maldives. This species can be distinguished from the only other member of the genus by the presence of more branched colonies, branches not being adnate to pedicels, longer pedicels, larger nematocysts, nematocyst-rich nematophore-like outgrowths from pedicels, smaller male gonophores, and a different geographic distribution. This finding represents the first record of the genus outside the type locality of its type species, in Belize. Molecular phylogenetic analyses show that, as expected, T. marhei belongs to the clade Filifera IV. However, the phylogenetic hypothesis based on both mitochondrial and nuclear DNA sequences reveals that most of the families of this group are polyphyletic, including Oceaniidae, and suggests that the morphological characters used to discriminate among filiferan families need to be revised thoroughly.

http://zoobank.org/urn:lsid:zoobank.org:pub:CE901E0D-B125-4A87-BB97-8020C0658B5D     

Assessing the efficacy of coherent anti‐Stokes Raman scattering microscopy for the detection of infiltrating glioblastoma in fresh brain samples

Coherent anti‐Stokes Raman scattering (CARS) microscopy is an emerging technique for identification of brain tumors. However, tumor identification by CARS microscopy on bulk samples and in vivo has been so far verified retrospectively on histological sections, which only provide a gross reference for the interpretation of CARS images without matching at cellular level. Therefore, fluorescent labels were exploited for direct assessment of the interpretation of CARS images of solid and infiltrative tumors. Glioblastoma cells expressing green fluorescent protein (GFP) were used for induction of tumors in mice (n = 7). The neoplastic nature of cells imaged by CARS microscopy was unequivocally verified by addressing two‐photon fluorescence of GFP on fresh brain slices and in vivo. In fresh unfixed biopsies of human glioblastoma (n = 10), the fluorescence of 5‐aminolevulinic acid‐induced protoporphyrin IX was used for identification of tumorous tissue. Distinctive morphological features of glioblastoma cells, i.e. larger nuclei, evident nuclear membrane and nucleolus, were identified in the CARS images of both mouse and human brain tumors. This approach demonstrates that the chemical contrast provided by CARS allows the localization of infiltrating tumor cells in fresh tissue and that the cell morphology in CARS images is useful for tumor recognition.

Experimental glioblastoma expressing green fluorescent protein.     

Soluble N‐ethylmaleimide‐sensitive factor attachment protein receptors required during Trypanosoma cruzi parasitophorous vacuole development

Trypanosoma cruzi, the etiologic agent of Chagas disease, is an obligate intracellular parasite that exploits different host vesicular pathways to invade the target cells. Vesicular and target soluble N‐ethylmaleimide‐sensitive factor attachment protein receptors (SNAREs) are key proteins of the intracellular membrane fusion machinery. During the early times of T. cruzi infection, several vesicles are attracted to the parasite contact sites in the plasma membrane. Fusion of these vesicles promotes the formation of the parasitic vacuole and parasite entry. In this work, we study the requirement and the nature of SNAREs involved in the fusion events that take place during T. cruzi infection. Our results show that inhibition of N‐ethylmaleimide‐sensitive factor protein, a protein required for SNARE complex disassembly, impairs T. cruzi infection. Both TI‐VAMP/VAMP7 and cellubrevin/VAMP3, two v‐SNAREs of the endocytic and exocytic pathways, are specifically recruited to the parasitophorous vacuole membrane in a synchronized manner but, although VAMP3 is acquired earlier than VAMP7, impairment of VAMP3 by tetanus neurotoxin fails to reduce T. cruzi infection. In contrast, reduction of VAMP7 activity by expression of VAMP7's longin domain, depletion by small interfering RNA or knockout, significantly decreases T. cruzi infection susceptibility as a result of a minor acquisition of lysosomal components to the parasitic vacuole. In addition, overexpression of the VAMP7 partner Vti1b increases the infection, whereas expression of a KIF5 kinesin mutant reduces VAMP7 recruitment to vacuole and, concomitantly, T. cruzi infection. Altogether, these data support a key role of TI‐VAMP/VAMP7 in the fusion events that culminate in the T. cruzi parasitophorous vacuole development.